The overall goal of these studies is to develop a novel adenovirus-lentivirus hybrid vector system for efficient gene delivery and long-term gene expression, and to use this hybrid vector as a vehicle for stable transduction of therapeutic genes for cancer gene therapy. To this end, we will construct and test helper-dependent adenovirus vectors that contain a complete packaging system for production of lentivirus vectors in situ. We will first test the hybrid vectors in cell culture studies to optimize the vector-associated parameters that might affect in situ second-stage lentivirus vector production and subsequent stable transduction efficiency. To explore the potential of these hybrid vectors for use in therapeutic applications, we will test their ability to achieve efficient in vivo gene delivery and long-term expression of an anti-angiogenic factor, endostatin, in an animal model of prostate cancer. Although previous studies have demonstrated tumor inhibition by endostatin, significant problems with systemic administration of this peptide have been encountered, including difficulty in producing adequate amounts of functional protein for in vivo use, and instability resulting in a very short circulating half-life, which necessitates high-dose bolus or continuous infusion schedules to maintain effective serum concentrations. We hypothesize that the adenovirus-lentivirus hybrid vector system has the potential to achieve high-level and long-term anti-angiogenic gene expression in vivo, and may thereby reduce local tumor growth and metastatic potential after intra-tumoral administration. We will first develop adenovirus-lentivirus hybrid vectors for stable gene expression, and evaluate the therapeutic efficacy of the anti-angiogenic gene in vitro. Subsequently, by using a mouse model of prostate cancer, we will evaluate the therapeutic efficacy of adenovirus--lentivirus hybrid-mediated gene transfer after intra-tumoral administration. These studies should ultimately lead to development of more efficient and widely applicable cancer gene therapy protocols. PROPOSED COMMERCIAL APPLICATION: Highly efficient and permanent delivery of therapeutic genes for cancer treatment or gene replacement therapy.